Steroid estrogen sulfates and methods of preparing the same



United States Patent ABSTRACT OF THE DISCLOSURE The preparation ofaromatic steroid triethylammonium sulfate salts by reacting astradiol orestriol with sulfur trioxide-triethylamine in an anhydrous solvent, isdescribed. These steroid salts are useful as estrogenic agents.

DESCRIPTION OF THE INVENTION This invention relates to new steroidcompounds. More particularly, it relates to aromatic steroidtriethylammonium sulfate salts and methods of preparing the same.

The novel steroid salts of this invention may be illustrated by thefollowing formula:

wherein X-Y is selected from the group consisting of The steroid saltsof the present invention are solids having well defined melting points.They are more water soluble than the steroids from which they areprepared, such as estradiol and estriol.

The compounds of the present invention are prepared by several methodsfor example, by reacting estradiol or estriol with sulfurtrioxide-triethylamine in an anhydrous solvent such as pyridine. Thereaction will take place at a temperature within the range of from to100 C. over a period of from 10 minutes to 24 hours. The major productobtained when estradiol is used is the 17fi-sulfooxy triethylammoniumsalt and when estriol is used, the 160:- sulfooxy triethylammonium salt.

The present compounds can also be prepared by a fusion process usingestradiol or estriol with sulfur trioxide-triethylamine. The fusionprocess is carried out at a temperature of 160 C. to 250 C. for a periodof from 5 minutes to minutes. The products obtained are identical withthose obtained when the reaction is carried out in an anhydrous solvent.

A still further method of preparing the present compounds is the fusionreaction of an estriol-3-sulfate triethylammonium salt orestradiol-3-sulfate triethylammonium salt. This reaction is carried outby merely heating the compounds at a temperature of from 140 C. to 250C. for from 5 minutes to 20 minutes. The fusion causes the shift of the3-sulfate triethylammonium salt to the 3,549,674 Patented Dec. 22, 197017-p0sition in the estradiol derivative, and to the 16aposition in theestriol derivative. The fusion reaction can also be carried out wherein17 3-estradio1 and 3-sulfooxyestra-1,3,5(l0)-triene triethylammoniumsalt are fused to produce l7fi-sulfooxyestra-1,3,5(l0)-trien-3-oltriethylammonium salt. v

The present compounds can also be obtained by removal of a 3-loweralkanoyl group from compounds, such as for example,S-acetoxy-17B-sulfooxyestra-1,3,5(10)- triene triethylamine,B-acetoxy-l6a-sulfooxyestra 1,3, 5(10)-trien-17,8-ol triethylamine, andthe like, by heating with triethylamine in the presence of solvents suchas alcohols. This hydrolysis method produces the present compoundswithout affecting the triethylammonium sulfate groups present.

The present compounds are useful as estrogenic agents. They may be usedin estrogen replacement therapy in the form of tablets, capsules, pills,solutions and the like. They can also be used parenterally in the formof sterile solutions or suspensions.

DETAILED DESCRIPTION The following examples describe in detail thepreparation of representative compounds of this invention.

EXAMPLE 1 Preparation of 17B-sulfooxyestra-l,3,5(l0)-trien-3-oltriethylammonium salt (a) From 17B-estradiol in pyridine solvent:

To a solution of estra-1,3,5(10)-triene-3,17B-diol (2.0 g.) in drypyridine (12 ml.) is added recrystallized sulfur trioxide-triethylamine(1.47 g., 1.1 eq.). After two hours at room temperature, the reactionmixture is poured into anhydrous ether, and an oil separates. Thesolution is decanted, more anhydrous ether is added and this processrepeated once again yielding a solid (2.02 g.). Thin layerchromatographic analysis indicated some 17,Bestradiol, a trace ofdisulfate but mostly a monosulfate. The solid is dissolved in methylenechloride and passed through a short pad of hydrous magnesium silicatewith additional methylene chloride (400 ml). The gradual addition ofanhydrous ether to the refluxing methylene chloride solution results inthe precipitation of the salt (0.8 g.). Additional quantities of thismaterial is obtained by further concentration of the mother liquor. Theanalytical sample melts at 200-20l C.

(b) From Uri-estradiol by fusion: A mixture of estra- 1,3,5 (10) triene3,175 diol (2.0 g.) and recrystallized sulfur trioxide-triethylamine(1.45 g.) are mixed together in a test tube. The test tube is insertedinto an oil bath whose temperature is C. The tube is kept in the bathapproximately 5 minutes by which time the temperature of the bath hasreached 180 C. The contents of the tube melts to a clear melt and thenresolidifies (in bath). The test tube is cooled and methylene chlorideis added to 3 dissolve the reaction mixture. In order to get a one phasesystem the final volume of methylene chloride is increased to 125milliliters. Refluxing this solution with the addition of anhydrousether gives an initial crystalline precipitation of product (1.70 g.),melting point 198200 C. Three subsequent concentrations of the motherliquor precipitates an additional amount (1.52 g.) of comparable qualitymaterial.

(c) From 17/8-estradiol by fusion:

OH I J 636 2Ha)aNHS 01-O- To 15 ml. of 88% formic acid there is added1.0 g. of 17,3-estradiol. The solution is heated on a steam-bath for onhour and is evaporated under reduced pressure to provide a gum.Methylene chloride is added to the gum and then removed under reducedpressure several times and finally anhydrous ether is added to theresidue whereupon the gum crystallized. The solid is collected and driedto give 1.0 g. of 17,8-estradiol-l7-formate. A sample is removed forrecrystallization from methylene chloride-hexane, melting point 154-155C.

In 1 ml. of pyridine there is dissolved 400 mg. of 17B-estradiol-l7-formate and 288 mg. of triethylamine-sulfur trioxide. Thesolution is allowed to remain overnight at room temperature, and is thenpoured into ether. The resulting gum, estradiol-3-triethylammoniumsulfate-17- formate, did not readily crystallize, and is dissolved in5.0 ml. of methanol and 0.5 ml. of triethylamine. The solution isrefluxed for hours and evaporated under reduced pressure to leave a gum(270 mg.) which crystallized on trituration with an hydrous ether.Recrystallization from methylene chloride-ether gives the desiredcompound, melting point 148150 C.

3-SulfooXyestra-l,3,5(10) trien 175-01 triethylammonium salt (101 mg.)is heated in a test tube at 140-170 C. over a period of minutes. Thematerial first melts and then resolidifies. The cooled solid shows aminor trace of estra 1,3,5(10) triene 3,175 diol on thin layerchromatography analysis but mostly a monosulfate, melting point -192 C.Recrystallization of this material from methylene chloride-anhydrousether gives the 17- sulfate (58 mg), melting point 198200 C., identicalto an authentic sample.

(e) From 17fi-estradiol-3-acetate-l7fl-triethylammoniurn salt:

Methanol (10.0 ml.), triethylamine (1.0 ml.) and 3- acetoxy-l7fisulfooxyestra 1,3,5( 10) triene triethylam monium salt (0.10 g.) arerefluxed for 0.5 hour and then the solvents are removed under reducedpressure. Crystallization of the residue from methylenechlorideanhydrous ether gives a solid (0.036 g.). The infrared spectrumof this material indicates a trace of 3-acetate remaining. Thehydrolysis is repeated using this solid and methanol 10.0 ml.) andtriethylamine (1.0 ml.) by refluxing for 45 minutes. Evaporation of thesolvents and recrystallization as above gives the l7-sulfate (0.030 g.),melting point 200201 C.

EXAMPLE 2 Preparation of 3 acetoxy 17,6 sulfooxyestra-1,3,5(10)- trienetriethylammonium salt (a) From 17,8-estradiol:

JOH

The sulfation of estra-1,3,5()-triene-3,17[3-diol (2.0 g.) in drypyridine (12 ml.) with sulfur trioxide-triethylamine (1.47 g.) isallowed to proceed for 2 hours at room temperature and then aceticanhydride (1.0 ml.) is added. After standing overnight at roomtemperature the reaction mixture is poured into anhydrous ether to givea solid which is filtered and dried. Solution of thesolid in methylenechloride and passage through a short pad of hydrous magnesium silicatefollowed by an additional 200 ml. of solvent afforded a filtrate whichdeposits the desired 3- acetate-17-sulfate (2.50 g., melting point166167 C.) on the addition of anhydrous ether. The addition of moreether afforded additional material (0.22 g.) melting point 165 166 C.Recrystallization from methylene chlorideanhydrous ether gives ananalytical sample (2.02 g.), melting point 17l172 C.

The stepwise addition of the two reagents, sulfur trioxide-triethylamineand acetic anhydride is not required to obtain the 3-acetate-l7-sulfatesince in a subsequent experiment when estradiol, in pyridine and the tworeagents in pyridine were combined, an equally good yield of the17-sulfate-3-acetate is obtained.

EXAMPLE 3 Preparation of 3 acetoxy-17fl-sulfooxyestra-1,3,5 10)triene-triethylammonium salt volume of methylene chloride and passedthrough a short hydrous magnesium silicate pad with additional methylenechloride passed through. Evaporation of the total filtrate andrecrystallization of the solid from hexane gives the 3- monoacetate(1.02 g.), melting point l42143 C.

Reported melting point 136.5137.5 (K. Miescher and C. Scholz, Helv.Chim. Acta, 20, 263 (1937)); reported melting point 14l142, [a] F58(dioxane),

ethanol max.

269 and 275 My. (e 768, 762) [H. Fex, K-E Lundvall, and A. Olsson, Acta.Chem. Scand. 22, 254 1968)].

After remaining 3 hours at room temperature, a reaction mixture of3-acetoxyestna-1,3,5(10)-trien-17I3-ol (0.75 g.), sulfurtrioxide-triethylamine (0.53 g.) in dry pyridine (2.0 ml.) is pouredinto anhydrous ether. The solid collected on filtration is dissolved ina minimum EXAMPLE -4 Preparation of 16a-sulfooxyestra-1,3,5(10)-triene-3-17B- diol triethylammonium salt (a) From estriol:

GHa

OH i "05%3 $11- A solution of estriol,[estra-1,3,5(10)-triene-3,16a-17/3- triol] (0.5 g.) in dry pyridine (10ml.) is stirred rapidly and a solution of sulfur trioxide-triethylamine(0.314 g.) in dry pyridine (5 ml.) is added dropwise over a period of 30minutes. The stirring is continued for 1 hour longer, and then thereaction mixture is poured into anhydrous ether giving an oily residue.The solvents are decanted and additional anhydrous ether is added. Afterbeing decanted again, and treated with additional ether, this materialis refrigerated overnight. Removal of the ether gives a solid which whenexamined by thin layer chromatography showed mostly monosulfate withtrace quantities of starting material and disulfate. Crystallizationfrom methanol-anhydrous ether removed the trace impurities, and thematerial (0.14 g.) melting point -151 C. is recrystallized to yield ananalytical sample (0.08 g.) melting point -163 C.

(b) From estriol by fusion:

fiuxed and anhydrous ether is added to turbidity. On cooling, acrystalline precipitate (0.205 g.), melting point l65167 C. is obtained.The identity of this material is confirmed by the infrared and nmranalysis.

(c) From estriol-3-sulfate triethylammonium salt by fusion:

[a] 22.7 (CHCl was 282 and 288 mp. (e 2160 and 1890 respectively).

A solution of estriol-16,17-diformate (0.5 g.) and triethylamine-sulfurtrioxide complex [0.29 g. (1.1 equiv.)] in dry pyridine (2.0 ml.) isallowed to stand overnight at room temperature. On pouring intoanhydrous ether, an oil is obtained from which the solvent is decanted.The oil is dissolved in methylene chloride and passed through a shortpad of hydrous magnesium silicate, and the resultant methylene chloridesolution is evaporated to give 0.41 g. o'f triethylammoniumestriol-l6,17-diformyl-2-sulfate as a glass.

The glass is refluxed for 1 hour in a solution of methanol ml.) andtriethylamine (1.0 ml.), and then the entire reaction mixture isevaporated to give a crystalline residue (0.35 g.). Severalrecrystallizations from acetonehexane gives the desired product (0.15g.), melting point 137138 C., [a] +28.1 (CHC1 silicate and the padfurther washed with a solution of I acetone (4 ml.)-methylene chloride(12 ml.). The combined eluate is refluxed and anhydrous ether added. Thecompound oiled from solution and then slowly crystallizes (0.11 g.),when a seed crystal is added, melting point 165168 C. The infraredspectrum of this material is identical to the previously preparedstandard.

EXAMPLE 5 Preparation of l7oc-SUlfOOXYeStl'fl-1,3,5(l0) trien-3-oltriethylammonium salt 9 6B CH3 osommonm;

To a solution of estra-1,3,S(l0)-triene-3,l7a-diol (0.5 g.) in drypyridine (1.5 m1.) is added recrystallized sulfur trioxidetriethylamine(0.37 g.). After remaining overnight at room temperature, the reactionmixture is poured into anhydrous ether, and an oil separates. Thesolution is decanted, more anhydrous ether is added and this processrepeated once again to yield a solid. The solid is dissolved inmethylene chloride and passed through a short pad of anhydrous magnesiumsilicate with additional methylene chloride passed through the absorbentpad. The combined effiuent is brought to reflux and anhydrous ether isgradually added to the point of turbidity. On cooling the desiredsulfate, triethylammonium salt slowly crystallizes (0.15 g.), meltingpoint 173-175 C.

What is claimed is:

1. A compound of the formula:

wherein X--Y is selected from the group consisting of wherein XY isselected from the group consisting of which comprises reacting acompound of the formula:

wherein -AB- is selected from the group consisting of I (1:112 (|3H--0Hwith sulfur trioxidetriethylamine in the presence of an anhydroussolvent at a temperature within the range of C. to 100 C. and recoveringsaid compounds therefrom.

6. A method in accordance with claim 5, in which the starting materialis estra-1,3,5(10)-triene-3,17/8-diol and the product obtained isl7fl-sulfooxyestra-l,3,5(10)-trien- 3-ol triethylammonium salt.

7. A method of preparing compounds o'f the formula:

wherein --X -Y is selected from the group consisting of which comprisesfusing a compound of the formula:

wherein -A -B is selected from the group consisting of with sulfurtrioxide triethylamine at a temperature within the range of 160 C. to250 C. for from 5 to 30 minutes and recovering said compounds therefrom.

8. A method of preparing compounds of the formula:

wherein -X --Y is selected from the group consisting of 6 GB CH2 OH--OS03 NHwQI-ma which comprises heating a compound of the formula:

xlll

wherein -A --B is selected from the group consisting of UNITED STATESPATENTS 2,534,121 12/1950 Grant et al. 2,835,681 5/1958 Allais et al.

HENRY A. FRENCH, Primary Examiner US. Cl. X.R. 260-999

